Enzyme immobilization on tritylagarose

A method is described for the immobilization on tritylated agarose or Sepharose columns of a wide spectrum of enzymes, including types useful in contemporary biochemistry/molecular biology, many of which have never before been reported as immobilized. The method involves the formation of noncovalent hydrophobic bonds between the enzymes and trityl groups which are attached to the agarose by means of ether bonds. The immobilization of calf intestinal and E. coli alkaline phosphatases to tritylagarose is reported in detail. Their binding strength, binding capacity, and long-term stability (greater than six months) are described as a function of the salt concentration, pH, buffer type, and degree of agarose substitution. Homologies are noted between tritylagarose-bound and membrane-bound phosphatases. This method compares favorably with other methods, covalent or otherwise, reported to date, in terms of the enzyme immobilization yield (ca. 100%), the mildness of conditions, resulting, in most cases, in the retention of a high degree of activity, the ease and speed of the manipulations, and the long-term stability of the immobilized enzyme. Further, it is noted that highly tritylated and crosslinked Sephadex G10 selectively and mildly removes detergents from enzyme solutions.     

Methods for studying aquatic bacteriophage ecology

Methods are described for enumerating the different kinds of bacteriophage present in virus concentrates prepared from 120 liters of water. Although developed specifically for use with coliphages, they should be applicable to viruses infecting other hosts. These methods involve mixed indicators, equilibrium buoyant-density centrifugation, use of enzymes or inhibitors or both, and specific hybridization probes, either alone or in combination. With these methods, it was possible to specifically enumerate filamentous and isometric male-specific (F-specific) phages, the different classes of P-group plasmid-specific phages, phiX174-like phages, and lambda-like phages. Some applications of these methods, including measurement of virus inactivation in the environment and the extent of fecal pollution, are discussed.     

Mechanisms of oleoylethanolamide-induced changes in feeding behavior and motor activity

Oleoylethanolamide (OEA), a lipid synthesized in the intestine, reduces food intake and stimulates lipolysis through peroxisome proliferator-activated receptor-alpha. OEA also activates transient receptor potential vanilloid type 1 (TRPV1) in vitro. Because the anorexigenic effect of OEA is associated with delayed feeding onset and reduced locomotion, we examined whether intraperitoneal administration of OEA results in nonspecific behavioral effects that contribute to the anorexia in rats. Moreover, we determined whether circulating levels of other gut hormones are modulated by OEA and whether CCK is involved in OEA-induced anorexia. Our results indicate that OEA reduces food intake without causing a conditioned taste aversion or reducing sodium appetite. It also failed to induce a conditioned place aversion. However, OEA induced changes in posture and reduced spontaneous activity in the open field. This likely underlies the reduced heat expenditure and sodium consumption observed after OEA injection, which disappeared within 1 h. The effects of OEA on motor activity were similar to those of the TRPV1 agonist capsaicin and were also observed with the peroxisome proliferator-activated receptor-alpha agonist Wy-14643. Plasma levels of ghrelin, peptide YY, glucagon-like peptide 1, and apolipoprotein A-IV were not changed by OEA. Finally, antagonism of CCK-1 receptors did not affect OEA-induced anorexia. These results suggest that OEA suppresses feeding without causing visceral illness and that neither ghrelin, peptide YY, glucagon-like peptide 1, apolipoprotein A-IV, nor CCK plays a critical role in this effect. Despite that OEA-induced anorexia is unlikely to be due to impaired motor activity, our data raise a cautionary note in how specific behavioral and metabolic effects of OEA should be interpreted.     

Honey bee colonies are group-level adaptive units

It is not widely recognized that natural selection has produced adaptive units at the level of groups. Multilevel selection theory shows that groups can evolve a high level of functional organization when between-group selection predominates over within-group selection. Strong empirical evidence that natural selection has produced adaptive units at the group level comes from studies of social insects in which we find colonies in certain species functioning as highly integrated units. The functional organization of a social insect colony is best understood for honey bees. Recent experimental analyses of honey bee colonies have revealed striking group-level adaptations that improve the foraging efficiency of colonies, including special systems of communication and feedback control. These findings are reviewed with the aim of showing that evolution has produced adaptively organized entities at the group level.